OLIGONUCLEOTIDE INHIBITION OF CELL PROLIFERATION

描述：针对C-myc癌基因的反义DNA阻断肿瘤 正常小鼠的生长。寡核苷酸的首次临床试验 治疗剂已经在患者中证明了反义功效， 滤泡性淋巴瘤、慢性髓细胞性白血病、巨细胞病毒和 克罗恩病。本申请的中心假设是， 而第三代ant-c-myc DNA的衍生物将显示出更大的 在c-myc的动物模型中， 依赖性淋巴瘤和其它依赖myc的恶性肿瘤。以前的工作 在这项资助下，已经证明了各种DNA的功效， 在恶性细胞中针对各种c-myc mRNA靶点的衍生物 在培养物和鼠宿主中生长。在接下来的五年里，这些 成功将转化为先导化合物。1.更敏感的c-myc 将识别mRNA靶点。放射性标记的c-myc mRNA靶点将被 与固定在 聚丙烯切片。为了测试杂交结果， 将测量一系列重叠的抗c-myc寡聚体的功效 在培养的淋巴瘤细胞中。2.抗c-myc DNA的效力将是 增加对于1.中确定的最佳序列，抗c-myc DNA 嵌合体将与末端硫代磷酸酯和中心硫代磷酸酯一起合成。 甲基膦酸酯或2 ′-O-甲基残基。肽核酸 还将制备具有设计的环肽配体的衍生物， 用于受体特异性细胞摄取。对于每种先导化合物， 特异性和机制将用1-4个错配进行测试。适体 效果将被筛选。3.抗c-myc DNA的药效学将 优化。(a)来自Eu-myc淋巴瘤细胞的肿瘤细胞，移植 进入免疫相容的亲本菌株，将用最多的 有效的抗c-myc DNA衍生物。肿瘤大小、mRNA和抗原 水平将作为剂量和时间表的函数进行研究。（B） 腹膜内、皮下、皮下贮库和口服给药 将被比较。(c)最有效的抗c-myc DNA将被测试， 残留疾病的辅助治疗，通过允许移植淋巴瘤 在开始治疗之前，细胞建立不同的时间。4. 将评价抗c-myc DNA的安全性。(a)行政、 新的抗c-myc衍生物的分布、代谢和排泄将 进行比较，以阐明效价随途径的变化。(b)体重， 脾肿大、细胞因子、补体、肝酶和全血 将比较计数以鉴定抗c-myc衍生物与 最大的治疗剂量5.抗c-myc DNA的谱宽 将被确定。将检测候选序列的疗效 针对人（a）ST 486伯基特淋巴瘤细胞（B）U937的异种移植物 红白血病细胞（c）BT474乳腺癌细胞（d）COLO 320结肠癌细胞 癌细胞和（e）A549肺癌细胞。的功效和效力 将评价联合治疗。抗c-myc DNA将在 与（a）免疫刺激性DNA或（B）抗- 增殖性化合物这些研究中产生的先导化合物 然后将被转换为一个单独的协作应用程序， I/II期临床试验。

DESCRIPTION: Antisense DNA directed against C-myc oncogene blocks tumor growth in normal mice. The first clinical trials of oligonucleotide therapeutics have demonstrated antisense efficacy in patients against follicular lymphoma, chronic myelogenous leukemia, cytomegalovirus and Crohn's disease. The central hypothesis of this application is that second and third generation derivatives of ant-c-myc DNAs will display greater systemic potency than phosphorothioates in animal models of c-myc- dependent lymphomas and other myc dependent malignancies. Previous work under this grant has demonstrated efficacy of a variety of DNA derivatives, against a variety of c-myc mRNA targets, in malignant cells growing in culture and in murine hosts. Over the next five years, these successes will be translated into lead compounds. 1. More sensitive c-myc mRNA targets will be identified. Radio-labeled c-myc mRNA targets will be hybridized to overlapping antisense DNA oligomers immobilized on polypropylene chips. To test the hybridization results, the antisense efficacy of a series of overlapping anti-c-myc oligomers will be measured in cultured lymphoma cells. 2. The potency of anti-c-myc DNAs will be increased. For the best sequences identified in 1., anti-c-myc DNA chimeras will be synthesized with terminal phosphorothioates and central methylphosphonates, or 2'-O-methyl residues. Peptide nucleic acid derivatives will also be prepared, with cyclic peptide ligands designed for receptor specific cellular uptake. For each lead compound, sequence specificity and mechanism will be tested with 1-4 mismatches. Aptameric effects will be screened. 3. The pharmacodynamics of anti-c-myc DNAs will be optimized. (a) Tumor cells from Eu-myc lymphoma cells, transplanted into the immuno-compatible parental strain, will be treated with the most potent anti-c-myc DNA derivatives above. Tumor size, mRNA, and antigen levels will be studied as a function of dose and schedule. (b) Intraperitoneal, subcutaneous, subcutaneous depot, and oral administration will be compared. (c) The most potent anti-c-myc DNAs will be tested for adjuvant therapy of residual disease, by allowing transplanted lymphoma cells to establish for varying times prior to initiation of therapy. 4. The safety of anti-c-myc DNAs will be evaluated. (a) Administration, distribution, metabolism, and excretion of new anti-c-myc derivatives will be compared to elucidate variation of potency with route. (b) Body weight, splenomegaly, cytokines, complement, liver enzymes and complete blood count will be compared to identify the anti-c-myc derivative with the greatest therapeutic ration. 5. The breadth of spectrum of anti-c-myc DNAs will be determined. Candidate sequences will be tested for efficacy against xenografts of human (a) ST486 Burkitt's lymphoma cells (b) U937 erythroleukemia cells (c) BT474 breast cancer cells (d) COLO320 colon cancer cells and (e) A549 lung cancer cells. The efficacy and potency of combination therapy will be evaluated. Anti-c-myc DNA's will be tested in combination or alteratively with (a) immunostimulatory DNAs or (b) anti- proliferative compounds. The lead compounds resulting from these studies will then be translated to a separate collaborative application for a Phase I/II clinical trial.